PROJECT SUMMARY/ABSTRACT 1 This proposal utilizes cutting-edge mechanistic approaches to investigate the clinically meaningful but hard-to- 2 study question of HOW insulin in human milk affects infant development. This research question is particularly 3 relevant to the growing and under-studied population of infants receiving breast milk from obese and/or insulin 4 resistant (IR) mothers - who produce milk with significantly greater insulin concentrations. Animal models show 5 that oral insulin delivered to the infant via breast milk has far-reaching effects on normal offspring development 6 and maturation. This proposal documents these effects in human infants and investigates how effects may be 7 altered in the context of maternal IR - and thus sustained infant exposure to high concentrations of oral insulin. 8 Specifically, we examine systemic effects regulated by infant pancreatic function as well as local intestinal 9 effects. Importantly, we will document both acute effects during the neonatal period when organs are still 10 immature AND effects of more ?chronic? exposure. We will recruit 64 exclusively breastfed infants of both 11 normal weight/normoglycemic (NW; n=28) and IR (n=36) mothers and study them during the neonatal period 12 (2-4 weeks) and after more ?chronic? exposure to mother?s milk (5 months). Our aims are: 13 Aim 1 ? Cross-Sectional Study ? Investigate differences in metabolic development and intestinal maturation 14 between infants exclusively breastfed by NW vs IR mothers at two critical time points: 2-4 weeks and 5 15 months. We hypothesize that infants breastfed by IR mothers will exhibit: an altered metabolomic profile 16 indicating differences in carbohydrate handling and insulin signaling (H1.1); an altered microbial composition in 17 the intestinal microbiome (H1.2); increased relative transcript abundance of insulin-target genes in exfoliated 18 intestinal cells, indicating more active insulin signaling at the level of the enterocyte (H1.3); and decreased 19 intestinal permeability (a functional measure of intestinal maturation) at 2-4 weeks (H1.4). 20 Aim 2 ? Glycemic Study - Determine if consumption of human milk from an IR mother (with high concentrations 21 of insulin) is correlated with differences in endogenous pancreatic response at 2-4 weeks. We hypothesize that 22 receipt of human milk with high insulin will result in a lower endogenous pancreatic insulin response (H2.1). 23 Aim 3 ? Glucose Tolerance Study - Determine if chronic consumption of human milk from an IR mother (with 24 high concentrations of insulin) is correlated with pancreatic response to a glucose challenge at 5 months. We 25 hypothesize that chronic exposure to human milk from an IR mother will under-stimulate pancreatic ?-cells and 26 result in a dampened endogenous insulin response to a standard glucose challenge at 5 months (H3.1). 27 Our long term objective is to fully understand the impact of oral insulin in the development and postnatal 28 programming of the breastfed infant. As the prevalence of insulin resistance rises among mothers ? and as 29 these women?s offspring have increased risk of developing metabolic disease themselves ? this work has far- 30 reaching implications regarding how we counsel women and care for their infants to minimize disease risk.